The most promising therapeutics for addressing morbidities that arise from congenital, traumatic, or infectious pathology lie in the field of regenerative medicine. On the forefront of these therapeutics is the field of regenerative dentistry. As we turn to cell-based therapies for repair of missing teeth and damaged oral tissues, we must look to developmental biology to understand how nature grew these tissues in the first place. What we know about tooth development or odontogenesis has been almost exclusively derived from the mouse. While we have learned a great deal from this system, in contrast to humans and most other vertebrates, the mouse lacks a successional or replacement set of teeth. We know relatively little about how to replace the dentition. To discover how nature perfected the growth of a tooth in an adult organism, we turn to the de-novo continuously replacing dentition of Lake Malawi cichlid fishes. In specific aim 1, I will identify dental stem cll populations responsible for natural de-novo tooth replacement in a perpetually replacing vertebrate dentition. I will utilize protocols I have developed to identify expression profiles of factors implicated in the stem cell biology of other vertebrate models. In supporting research, I noted that cichlid teeth and taste buds share a common epithelial ribbon, as well as gene co-expression, during development and regeneration. I will employ in-situ hybridization (ISH) for mRNAs and miRNAs to uncover factors expressed in both tooth and/or taste bud development which may be involved in their continuous turn-over. I will then employ pulse-chase analysis to verify the location and character of putative stem cells in support of my ISH experiments. In aim 2, I will manipulate molecular signals that putatively control the regeneration of teeth and taste buds. In this aim, I will treat fishes with small molecule chemical agonists and antagonists of major developmental pathways expressed in putative stem cell niches. I will use antagomirs to disrupt the function of miRNAs analyzed in aim one that are expressed in the putative stem niches. I will subsequently assay changes in mRNA, miRNA, and pulse-chase data in those treatments resulting in disrupted tooth replacement phenotypes. These experiments will demonstrate the importance of key factors in dental regeneration. As we move into an era where the replacement of missing or damaged tissues has become not only a promising therapy, but a viable possibility, it is imperative that we appreciate how those tissues were formed in the first place. With a more global health impact, we seek to identify the epithelial and mesenchymal stem cells responsible for the natural replacement of an adult organ. Part of the NIDCR strategic plan over the next years is to facilitate reconstruction and regeneration of diseased or damaged oral and craniofacial tissues and organs through biological, bioengineering and biomaterials research approaches. These findings will advance our understanding of dental stem cells so that we may one day design therapies based on endogenous mechanisms of repair.